Christopher B. Wilson

The Toll-like receptor 2 is recruited to macrophage phagosomes and discriminates between pathogens

Macrophages orchestrate innate immunity by phagocytosing pathogens and coordinating inflammatory responses. Effective defence requires the host to discriminate between different pathogens. The specificity of innate immune recognition in Drosophila is mediated by the Toll family of receptors; Toll mediates anti-fungal responses, whereas 18-wheeler mediates anti-bacterial defence. A large number of Toll homologues have been identified in mammals, and Toll-like receptor 4 is critical in responses to Gram-negative bacteria. Here we show that Toll-like receptor 2 is recruited specifically to macrophage phagosomes containing yeast, and that a point mutation in the receptor abrogates inflammatory responses to yeast and Gram-positive bacteria, but not to Gram-negative bacteria. Thus, during the phagocytosis of pathogens, two classes of innate immune receptors cooperate to mediate host defence: phagocytic receptors, such as the mannose receptor, signal particle internalization, and the Toll-like receptors sample the contents of the vacuole and trigger an inflammatory response appropriate to defence against the specific organism.

Regulation of interferon-gamma during innate and adaptive immune responses.

Interferon-gamma (IFN-gamma) is crucial for immunity against intracellular pathogens and for tumor control. However, aberrant IFN-gamma expression has been associated with a number of autoinflammatory and autoimmune diseases. This cytokine is produced predominantly by natural killer (NK) and natural killer T (NKT) cells as part of the innate immune response, and by Th1 CD4 and CD8 cytotoxic T lymphocyte (CTL) effector T cells once antigen-specific immunity develops. Herein, we briefly review the functions of IFN-gamma, the cells that produce it, the cell extrinsic signals that induce its production and influence the differentiation of naïve T cells into IFN-gamma-producing effector T cells, and the signaling pathways and transcription factors that facilitate, induce, or repress production of this cytokine. We then review and discuss recent insights regarding the molecular regulation of IFN-gamma, focusing on work that has led to the identification and characterization of distal regulatory elements and epigenetic modifications with the IFN-gamma locus (Ifng) that govern its expression. The epigenetic modifications and three-dimensional structure of the Ifng locus in naive CD4 T cells, and the modifications they undergo as these cells differentiate into effector T cells, suggest a model whereby the chromatin architecture of Ifng is poised to facilitate either rapid opening or silencing during Th1 or Th2 differentiation, respectively.

A Critical Role for Dnmt1 and DNA Methylation in T Cell Development, Function, and Survival

The role of DNA methylation and of the maintenance DNA methyltransferase Dnmt1 in the epigenetic regulation of developmental stage- and cell lineage-specific gene expression in vivo is uncertain. This is addressed here through the generation of mice in which Dnmt1 was inactivated by Cre/loxP-mediated deletion at sequential stages of T cell development. Deletion of Dnmt1 in early double-negative thymocytes led to impaired survival of TCRalphabeta(+) cells and the generation of atypical CD8(+)TCRgammadelta(+) cells. Deletion of Dnmt1 in double-positive thymocytes impaired activation-induced proliferation but differentially enhanced cytokine mRNA expression by naive peripheral T cells. We conclude that Dnmt1 and DNA methylation are required for the proper expression of certain genes that define fate and determine function in T cells.

Loss of genomic methylation causes p53-dependent apoptosis and epigenetic deregulation

Cytosine methylation of mammalian DNA is essential for the proper epigenetic regulation of gene expression and maintenance of genomic integrity. To define the mechanism through which demethylated cells die, and to establish a paradigm for identifying genes regulated by DNA methylation, we have generated mice with a conditional allele for the maintenance DNA methyltransferase gene Dnmt1. Cre-mediated deletion of Dnmt1 causes demethylation of cultured fibroblasts and a uniform p53-dependent cell death. Mutational inactivation of Trp53 partially rescues the demethylated fibroblasts for up to five population doublings in culture. Oligonucleotide microarray analysis showed that up to 10% of genes are aberrantly expressed in demethylated fibroblasts. Our results demonstrate that loss of Dnmt1 causes cell-type–specific changes in gene expression that impinge on several pathways, including expression of imprinted genes, cell-cycle control, growth factor/receptor signal transduction and mobilization of retroelements.

Epigenetic control of T-helper-cell differentiation

Naive CD4+ T cells give rise to T-helper-cell subsets with functions that are tailored to their respective roles in host defence. The specification of T-helper-cell subsets is controlled by networks of lineage-specifying transcription factors, which bind to regulatory elements in genes that encode cytokines and other transcription factors. The nuclear context in which these transcription factors act is affected by epigenetic processes, which allow programmes of gene expression to be inherited by progeny cells that at the same time retain the potential for change in response to altered environmental signals. In this Review, we describe these epigenetic processes and discuss how they collaborate to govern the fate and function of T helper cells.

Human Toll-like receptor 4 recognizes host-specific LPS modifications

Lipopolysaccharide (LPS) is the principal proinflammatory component of the Gram-negative bacterial envelope and is recognized by the Toll-like receptor 4 (TLR4)–MD-2 receptor complex. Bacteria can alter the acylation state of their LPS in response to environmental changes. One opportunistic bacterium, Pseudomonas aeruginosa, synthesizes more highly acylated (hexa-acylated) LPS structures during adaptation to the cystic fibrosis airway. Here we show that human, but not murine, TLR4–MD-2 recognizes this adaptation and transmits robust proinflammatory signals in response to hexa-acylated but not penta-acylated LPS from P. aeruginosa. Whereas responses to lipidIVA and taxol are dependent on murine MD-2, discrimination of P. aeruginosa LPS structures is mediated by an 82-amino-acid region of human TLR4 that is hypervariable across species. Thus, in contrast to mice, humans use TLR4 to recognize a molecular signature of bacterial-host adaptation to modulate the innate immune response.

Cutting Edge: Functional Interactions Between Toll-Like Receptor (TLR) 2 and TLR1 or TLR6 in Response to Phenol-Soluble Modulin

Toll-like receptor (TLR) 2 and TLR4 play important roles in the early, innate immune response to microbial challenge. TLR2 is preferentially involved in the inflammatory response to lipoteichoic acid, lipopeptides, and glycans from a variety of microbes, whereas TLR4 is essential for a complete response to LPSs. We report here that TLR2 transduces the response to phenol-soluble modulin, a factor secreted by Staphylococcus epidermidis. The TLR2-mediated response to this modulin was enhanced by TLR6 but inhibited by TLR1, indicating a functional interaction between these receptors. We also demonstrate that a response to phenol-soluble modulin mediated by TLR2 and TLR6 was more refractory to inhibition by TLR1 than one mediated by TLR2 alone.

Neonatal Innate TLR-Mediated Responses Are Distinct from Those of Adults

The human neonate and infant are unduly susceptible to infection with a wide variety of microbes. This susceptibility is thought to reflect differences from adults in innate and adaptive immunity, but the nature of these differences is incompletely characterized. The innate immune response directs the subsequent adaptive immune response after integrating information from TLRs and other environmental sensors. We set out to provide a comprehensive analysis defining differences in response to TLR ligation between human neonates and adults. In response to most TLR ligands, neonatal innate immune cells, including monocytes and conventional and plasmacytoid dendritic cells produced less IL-12p70 and IFN-α (and consequently induced less IFN-γ), moderately less TNF-α, but as much or even more IL-1β, IL-6, IL-23, and IL-10 than adult cells. At the single-cell level, neonatal innate cells generally were less capable of producing multiple cytokines simultaneously, i.e., were less polyfunctional. Overall, our data suggest a robust if not enhanced capacity of the neonate vs the adult white-blood cell TLR-mediated response to support Th17- and Th2-type immunity, which promotes defense against extracellular pathogens, but a reduced capacity to support Th1-type responses, which promote defense against intracellular pathogens.

Immunologic basis for increased susceptibility of the neonate to infection

Neonatal infection with the GBS occurs in a small fraction of exposed infants who lack specific antibody. Diminished influx of PMNs to sites of infection as a result of abnormalities in chemotaxis, bone marrow exhaustion, and to a lesser degree relative complement deficiency and decreased microbicidal activity of PMNs may be additional predisposing factors. Infection with HSV occurs more often in infants born to mothers with primary rather than secondary infection; the lack of passively acquired antibody in such infants is a possible but unproved susceptibility factor. The failure of neonates to control HSV may also be related to decreased production of or response to interferon or to decreased activity of nonimmune and immune cellular cytotoxic mechanisms. Similarly, infection with Toxoplasma and intracellular bacterial pathogens, such as Listeria, may be more severe because of the decreased generation of lymphokines and interleukins, which attract macrophages to the site of infection and enable them to kill these organisms. Much of this analysis based on in vitro and animal studies summarizes current information in a rapidly changing field rather than stating established fact. The precise age at which most of the immune functions discussed reach maturity is unknown. However, the risk of severe infection with these pathogens appears to wane by 2 to 3 months of age. Although this may partly reflect decreased exposure, we might hypothesize that immune functions that are mature by this age are those most critical for protection. Future studies focusing on changes in immune function during the first months of life may provide useful insights into the immunobiology of these diseases and direct attention to the most fruitful areas for immunologic intervention.

Inactivation of Notch1 in immature thymocytes does not perturb CD4 or CD8 T cell development

Notch proteins influence cell-fate decisions in many developing systems. Several gain-of-function studies have suggested a critical role for Notch1 signaling in CD4-CD8 lineage commitment, matura-tion and survival in the thymus. However, we show here that tissue-specific inactivation of the gene encoding Notch1 in immature (CD25+CD44−) T cell precursors does not affect subsequent thymocyte development. Neither steady-state numbers nor the rate of production of CD4+ and CD8+ mature thymocytes is perturbed in the absence of Notch1. In addition, Notch1-deficient thymocytes are normally sensitive to spontaneous or glucocorticoid-induced apoptosis. In contrast to earlier re-ports, these data formally exclude an essential role for Notch1 in CD4-CD8 lineage commitment, maturation or survival.